Design and characterisation of mutant and wild-type huntingtin proteins produced from a toolkit of scalable eukaryotic expression systems

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Harding, Rachel J. ; Loppnau, Peter ; Ackloo, Suzanne ; Lemak, Alexander ; Hutchinson, Ashley ; Hunt, Brittany ; Holehouse, Alex ; Ho, Jolene ; Fan, Lixin ; Toledo-Sherman, Leticia ; Seitova, Alma ; Arrowsmith, Cheryl (2018)
  • Related identifiers: doi: 10.5281/zenodo.2169035, doi: 10.5281/zenodo.2169034
  • Subject: Huntingtin | Huntington's disease | HD resources | Protein production
    mesheuropmc: nervous system diseases | congenital, hereditary, and neonatal diseases and abnormalities | nervous system | mental disorders

ABSTRACT: The pathogenic Huntington’s disease (HD) mutation causes polyglutamine (polyQ) tract expansion of the 348 kDa HTT protein above a critical threshold of ~35 glutamines. HD mutation effect on HTT is poorly understood, partly due to difficulties in performing biochemical studies with this large protein. To facilitate such studies, we generated resources for HTT production in multiple eukaryotic expression systems, comprising constructs with polyQ lengths representing general population, HD patients, juvenile HD patients and the more extreme expansions used in some tissue and animal models. These reagents yield milligram quantities of pure HTT protein. We biophysically characterised HTT samples produced using this HD resource, gleaning insight into the nature of full-length HTT in its apo form and when bound to its binding partner HAP40. Work outlined in this manuscript and the tools generated, lay a foundation for further biochemical study of the HTT protein and its functional interactions with other biomolecules.
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